A TRUNCATED FORM OF HEME OXYGENASE-1 IS FOUND IN THE NUCLEUS OF CULTURED CELLS IN RESPONSE TO INCREASES IN INTRACELLULAR HEME

Guang Yang, Ph.D.*, Qing Lin, Ph.D.*, Yi-Hao Weng, M.D.#. Kimberly Rish, M.S.**, Ann Smith, Ph.D. ** and Phyllis A. Dennery, M.D.

*†#.Children's Hospital of Philadelphia*, Department of Pediatrics, University of Pennsylvania†, Philadelphia, Pennsylvania; School of Biological Sciences, University of Missouri-Kansas City**, Kansas City, Missouri; and Department of Pediatrics, Stanford University#, California, U.S.A.

Heme oxygenase (HO)-1, the rate–limiting enzyme in heme degradation, is an integral membrane protein of the smooth endoplasmic reticulum. HO-1 is induced by various stimuli including oxidative stress, heavy metals, UV radiation and inflammation but also by heme and heme-hemopexin. Cytoprotective roles for HO-1 are established, although the mechanism by which this occurs is not yet fully defined. When mouse hepatoma cells are incubated either with exogenous heme or heme-hemopexin, a truncated form of HO-1 is detected in the nucleus that lacks the C-terminus.  Exogenous heme also generates a nuclear form of HO-1 in NIH 3T3 cells. Transfection of 3T3 cells with an N-terminal 6His-tagged/C-terminal FLAG-tagged HO-1 expression construct followed by incubation with heme leads to co-localization of the His, but not the FLAG signal, with a nuclear DAPI stain. Transfection with a GFP-HO-1 cDNA with a deletion of the C-terminus results in nuclear localization of the protein.  Deletion of a putative nuclear export sequence (NES) results in diffuse localization of HO-1 that is not enhanced by hemin. Thus, hemin-mediated modification of the NES is also required for nuclear translocation. This nuclear C-terminal truncated HO-1 lacks normal enzymic activity and may act as a transcriptional regulator.